CHOP (C/EBP homologous protein) and ASNS (asparagine synthetase) induction in cybrid cells harboring MELAS and NARP mitochondrial DNA mutations

Abstract

Mitochondrial dysfunction caused by mutations in mitochondrial DNA (mtDNA) is related to a variety of diseases including MELAS and NARP syndromes. However, little is known about the intracellular responses induced by mtDNA mutations. In order to identify genes whose expression is altered as a result of the presence of mtDNA mutations, DNA microarray analysis was performed using human 143B osteosarcoma cells harboring 3243A > G [tRNA-Leu (UUR)] and 8993T > G [ATPase6 Leu156Arg] mtDNA mutations associated with MELAS and NARP syndromes (2SD and NARP3-1 cybrid cells), respectively. We found that mRNA and protein levels of ATF4, CHOP and ASNS were upregulated in 2SD and NARP3-1 cells as compared with parental cells. Reporter assays demonstrated that transcription of CHOP and ASNS genes was upregulated through the AARE (amino acid regulatory element) and NSRE-1 (nutrient-sensing response element-1) enhancer elements to which ATF4 binds, respectively. Furthermore, knockdown of ATF4 by RNA interference reduced CHOP and ASNS transcription in 2SD and NARP3-1 cells. These results suggest that the presence of mtDNA mutations elicits upregulation of CHOP and ASNS genes through the elevation of ATF4 expression and its binding to the AARE and NSRE-1, respectively.